Method for forming nerve graft

ABSTRACT

A method for forming a nerve graft includes the following steps. A carbon nanotube structure is provided. A hydrophilic layer is formed on a surface of the carbon nanotube structure. The hydrophilic layer is polarized to form a polar surface on the hydrophilic layer. A number of neurons are formed on the polar surface of the hydrophilic layer to form a nerve network. The neurons connect with each other.

This application claims all benefits accruing under 35 U.S.C. §119 from China Patent Application No. 201110031597.2, filed on Jan. 28, 2011 and China Patent Application No. 201110101123.0, filed on Apr. 21, 2011 in the China Intellectual Property Office, disclosure of which is incorporated herein by reference.

CROSS-REFERENCES TO RELATED APPLICATIONS

This application is related to commonly-assigned applications entitled, “NERVE GRAFT,” filed Jan. 13, 2012 Ser. No. 13/349,582. “CULTURE MEDIUM,” filed Jan. 13, 2012 Ser. No. 13/349,656, and “METHOD FOR FORMING CULTURE MEDIUM,” filed Jan. 13, 2012 Ser. No. 13/349,660.

BACKGROUND

1. Technical Field

The present disclosure relates to a method for forming a nerve graft, especially to a method for forming a nerve graft for guiding injured neurons to reconnect.

2. Description of Related Art

A nervous system is a complex cellular communication network that is mainly composed of neurons and glial cells (neuroglial cells). Glial cells occupy spaces between the neurons and modulate the neurons' functions. The neurons sense stimuli and transmit this information to the brain for processing and storage. For example, the neurons receive diverse stimuli from the environment (e.g. light, touch, sound) and transmit electrical signals, which are then converted into chemical signals to be passed on to other cells.

Neurons exist in a number of different shapes and sizes, and can be classified by their morphology and function. The basic morphology of a neuron includes a cell body and neurites projecting/branching from the cell body towards other neurons. The neurites can also be divided into two types by their functions. One is a dendrite, which branches around the cell body and receive signals from other neurons to the cell body. The other is an axon, which branches from the cell body and grows continually without tapering. The axon conducts the signals away from the neuron's cell body. The end of the axon has branching terminals that release neurotransmitters into a gap between the branching terminals and the dendrites of other neurons. Thus, the information or signal is propagated.

Neuron damage can lead to neurite degeneration and retraction. If the damage is severe, breaks in neurites affect signal transmission and the cellular communication between neurons will cease.

What is needed, therefore, is a method for forming a nerve graft which can reconnect opposite terminals in broken neurites.

BRIEF DESCRIPTION OF THE DRAWINGS

Many aspects of the disclosure can be better understood with reference to the drawings. The components in the drawings are not necessarily drawn to scale, the emphasis instead being placed upon clearly illustrating the principles of the present disclosure. Moreover, in the drawings, like reference numerals designate corresponding parts throughout the several views.

FIG. 1 is a cross-sectional view of one embodiment of a nerve graft.

FIG. 2 shows a scanning electron microscope (SEM) image of a flocculated carbon nanotube film.

FIG. 3 shows an SEM image of a pressed carbon nanotube film.

FIG. 4 shows an SEM image of a drawn carbon nanotube film.

FIG. 5 shows an SEM image of a carbon nanotube structure.

FIG. 6 shows an SEM image of an untwisted carbon nanotube wire.

FIG. 7 shows an SEM image of a twisted carbon nanotube wire.

FIG. 8 is a top view of the nerve graft shown in FIG. 1.

FIG. 9 is a flowchart of one embodiment of a method for forming a nerve graft.

FIG. 10 shows an SEM image of a neuron cultured on a culture substrate of the nerve graft.

FIG. 11 shows an SEM image of the nerve graft shown in FIG. 1, wherein a nerve network of the nerve graft is dyed.

FIG. 12 is a cross-sectional view of another embodiment of a nerve graft.

FIG. 13 is a flowchart of another embodiment of a method for forming a nerve graft.

DETAILED DESCRIPTION

The disclosure is illustrated by way of example and not by way of limitation in the figures of the accompanying drawings in which like references indicate similar elements. It should be noted that references to “an” or “one” embodiment in this disclosure are not necessarily to the same embodiment, and such references mean at least one.

Referring to FIG. 1, one embodiment of a nerve graft 100 includes a culture substrate 10 and a nerve network 20 positioned on a surface of the culture substrate 10. The nerve network 20 includes a number of neurons 21. The adjacent neurons 21 connect with each other. The surface of the culture substrate 10 has a polarity which attracts a surface of the nerve network 20 such that the nerve network 20 can closely adhere to the surface of the culture substrate 10.

The culture substrate 10 includes a carbon nanotube structure 12 and a hydrophilic layer 14 located on a surface of the carbon nanotube structure 12. The hydrophilic layer 14 has a polar surface 16 located away from the carbon nanotube structure 12. The nerve network 20 is positioned on the polar surface 16 of the hydrophilic layer 14 such that the hydrophilic layer 14 is located between the carbon nanotube structure 12 and the nerve network 20.

The hydrophilic layer 14 is a hydrophilic environment for growing the neurons 21 of the nerve network 20. The thickness of the hydrophilic layer 14 is in a range from about 1 nanometer (nm) to about 100 nm. Preferably, the hydrophilic layer 14 has a thickness in a range from about 1 nm to about 50 nm. The hydrophilic layer 14 is made from inorganic materials, such as silicon dioxide, titanium dioxide, iron oxide, or any combination thereof. In one embodiment, the hydrophilic layer 14 is a silicon dioxide layer with a thickness of about 10 nm.

The polar surface 16 of the hydrophilic layer 14 has a polarity such that the polarity of the culture substrate 10 can attract the surface of the nerve network 20 with opposite affinity. Thus, the culture substrate 10 is capable of providing a bio-compatible environment for seeding the neurons 21 and forming the nerve network 20.

The carbon nanotube structure 12 is capable of forming a free-standing structure. The term “free-standing structure” can be defined as a structure that does not need to be supported by a substrate. For example, a free-standing structure can sustain the weight of itself if the free-standing structure is hoisted by a portion thereof without any significant damage to its structural integrity. Carbon nanotubes distributed in the carbon nanotube structure 12 defines a plurality of gaps therebetween. The carbon nanotubes can have a significant van der Waals attractive force therebetween. The free-standing structure of the carbon nanotube structure 12 is realized by the carbon nanotubes joined by van der Waals attractive force.

The carbon nanotubes in the carbon nanotube structure 12 can be orderly or disorderly arranged. The term ‘disordered carbon nanotube film structure’ includes, but is not limited to, a structure where the carbon nanotubes are arranged along many different directions such that the number of carbon nanotubes arranged along each different direction can be almost the same (e.g. uniformly disordered), and/or entangled with each other. The term ‘ordered carbon nanotube film structure’ includes, but is not limited to, a structure where the carbon nanotubes are arranged in a consistently systematic manner, e.g., the carbon nanotubes are arranged approximately along a same direction and or have two or more sections within each of which the carbon nanotubes are arranged approximately along a same direction (different sections can have different directions). The carbon nanotubes in the carbon nanotube structure 12 can be single-walled, double-walled, and/or multi-walled carbon nanotubes.

The carbon nanotube structure 12 can include a flocculated carbon nanotube film as shown in FIG. 2. The flocculated carbon nanotube film can include a number of long, curved, disordered carbon nanotubes entangled with each other and can form a free-standing structure. Furthermore, the flocculated carbon nanotube film can be isotropic. The carbon nanotubes can be substantially uniformly dispersed in the flocculated carbon nanotube film. The adjacent carbon nanotubes are acted upon by the van der Waals attractive force therebetween, thereby forming an entangled structure with micropores defined therein. Sizes of the micropores can be in a range from about 1 nm to about 500 nm. Due to the carbon nanotubes in the carbon nanotube structure 12 being entangled with each other, the carbon nanotube structure 12 employing the flocculated carbon nanotube film has excellent durability and can be fashioned into desired shapes with a low risk to the integrity of the carbon nanotube structure 12. The flocculated carbon nanotube film, in some embodiments, will not require the use of a structural support due to the carbon nanotubes being entangled and adhered together by van der Waals attractive force therebetween.

The carbon nanotube structure 12 can include a pressed carbon nanotube film. The carbon nanotubes in the pressed carbon nanotube film can be arranged along a same direction or arranged along different directions. The carbon nanotubes in the pressed carbon nanotube film can rest upon each other. The adjacent carbon nanotubes are combined and attracted to each other by van der Waals attractive force, and can form a free-standing structure. An angle between a primary alignment direction of the carbon nanotubes and a surface of the pressed carbon nanotube film can be in a range from about 0 degrees to about 15 degrees. The pressed carbon nanotube film can be formed by pressing a carbon nanotube array. The angle is closely related to pressure applied to the carbon nanotube array. The greater the pressure, the smaller the angle. The carbon nanotubes in the carbon nanotube film are substantially parallel to the surface of the carbon nanotube film if the angle is about 0 degrees. A length and a width of the carbon nanotube film can be set as desired. The pressed carbon nanotube film can include a number of carbon nanotubes substantially aligned along one or more directions. The pressed carbon nanotube film can be obtained by pressing the carbon nanotube array with a pressure head. Alternatively, the shape of the pressure head and the pressing direction can determine the direction of the carbon nanotubes arranged therein. Specifically, in one embodiment, a planar pressure head is used to press the carbon nanotube array along the direction substantially perpendicular to a substrate. A number of carbon nanotubes pressed by the planar pressure head may be sloped in many directions. In one embodiment, as shown in FIG. 3, if a roller-shaped pressure head is used to press the carbon nanotube array along a certain direction, the pressed carbon nanotube film having a number of carbon nanotubes substantially aligned along the certain direction can be obtained. In another embodiment, if the roller-shaped pressure head is used to press the carbon nanotube array along different directions, the pressed carbon nanotube film having a number of carbon nanotubes substantially aligned along different directions can be obtained.

In one embodiment, the carbon nanotube structure 12 includes at least one drawn carbon nanotube film as shown in FIG. 4. The drawn carbon nanotube film can have a thickness of about 0.5 nm to about 100 micrometers (∞m). The drawn carbon nanotube film includes a number of carbon nanotubes that can be arranged substantially parallel to a surface of the drawn carbon nanotube film. A plurality of micropores having a size of about 1 nm to about 500 nm can be defined by the carbon nanotubes. A large number of the carbon nanotubes in the drawn carbon nanotube film can be oriented along a preferred orientation, meaning that a large number of the carbon nanotubes in the drawn carbon nanotube film are arranged substantially along the same direction. An end of one carbon nanotube is joined to another end of an adjacent carbon nanotube arranged substantially along the same direction, by van der Waals attractive force. More specifically, the drawn carbon nanotube film includes a number of successively oriented carbon nanotube segments joined end-to-end by van der Waals attractive force therebetween. Each carbon nanotube segment includes a number of carbon nanotubes substantially parallel to each other and joined by van der Waals attractive force therebetween. The carbon nanotube segments can vary in width, thickness, uniformity, and shape. A small number of the carbon nanotubes are randomly arranged in the drawn carbon nanotube film and has a small if not negligible effect on the larger number of the carbon nanotubes in the drawn carbon nanotube film arranged substantially along the same direction.

In another embodiment, the carbon nanotube structure 12 can include a number of stacked drawn carbon nanotube films as shown in FIG. 5. Adjacent drawn carbon nanotube films can be adhered by the van der Waals attractive force therebetween. An angle can exist between the carbon nanotubes in adjacent drawn carbon nanotube films. The angle between the aligned directions of the adjacent drawn carbon nanotube films can be in a range from about 0 degrees to about 90 degrees. In one embodiment, the carbon nanotube structure 12 is formed by 30 layers of drawn carbon nanotube films. The angle between the aligned directions of the adjacent drawn carbon nanotube films is about 90 degrees. Simultaneously, aligned directions of adjacent drawn carbon nanotube films can be substantially perpendicular to each other, thus a plurality of micropores and nodes can be defined by the carbon nanotube structure 12.

Alternatively, the carbon nanotube structure 12 can be formed by a number of carbon nanotube wires. Thus, one portion of the carbon nanotube wires is arranged substantially parallel to each other and extends substantially along a first direction. In addition, the other portion of the carbon nanotube wires is arranged substantially parallel to each other and extends substantially along a second direction. The first direction and the second direction can be substantially perpendicular to each other. In one embodiment, the carbon nanotube wire can be classified as untwisted carbon nanotube wire and twisted carbon nanotube wire. Referring to FIG. 6, the untwisted carbon nanotube wire is made by treating an organic solvent to the carbon nanotube film described above. In such case, the carbon nanotubes of the untwisted carbon nanotube wire are substantially parallel to the axis of the carbon nanotube wire. In one embodiment, the organic solvent can be ethanol, methanol, acetone, dichloroethane, or chloroform. The diameter of the untwisted carbon nanotube wire is in a range from about 0.5 nm to about 1 millimeter.

Furthermore, referring to FIG. 7, the carbon nanotube wire can be formed by twisting the carbon nanotube film to form the twisted carbon nanotube wire. Specifically, twisted carbon nanotube wire is formed by turning two opposite ends of the carbon nanotube film in opposite directions. In one embodiment, the carbon nanotubes of the carbon nanotube wire are aligned around the axis of the carbon nanotube spirally.

Referring to FIG. 8, the nerve network 20 includes a number of neurons 21. Each neuron 21 includes a cell body 22 and at least one neurite 24 branching from the cell body 22. The neurites 24 of adjacent neurons 21 connect with each other such that the adjacent neurons 21 can connect with each other. The neurites 24 of the neurons 21 include dendrites and axons. Generally, the neurites 24 can grow in any direction from one cell body 22. However if there are a number of cell bodies 22 located on the surface of the culture substrate 10, the neurites 24 from one cell body 22 would preferentially extend to adjacent cell bodies 22. Thus, growth directions of the neurites 24 can be guided by positions of the cell bodies 22. In one embodiment, the neurons 21 are hippocampal neurons grown on the surface of the culture substrate 10 including 30 layers of carbon nanotube films. Specifically, the culture substrate 10 includes a silicon dioxide layer and 30 layers of carbon nanotube films. The hippocampal neurons are grown on a surface of the silicon dioxide layer.

The culture substrate 10 can define a growth surrounding the neurons 21, thus the nerve network 20 can be formed on the culture substrate 10. Both of the hydrophilic layer 14 and the carbon nanotube structure 12 can have good tactility, and nonmetal and bio-compatible properties. Thus, the culture substrate 10 including the hydrophilic layer 14 and the carbon nanotube structure 12 can be transplanted into a biological body and form a desired shape. Therefore, the shape and a thickness of the culture substrate 10 can be designed as a shape and a thickness of a wound on the biological body. The neurons 21 of the nerve network 20 can communicate with each other, so that if the nerve graft 100 is transplanted into the wound, neurons of the biological body close to the wound will communicate with and connect to the nerve network 20 of the nerve graft 100. Thus, the injured neurons can be reconnected together. An area of the wound can be substantially equal to an area of a cross-section of the nerve graft, and a distance between an edge of the nerve graft and an edge of the wound can be less than a length of the wound. Therefore, a distance between the neurons of the biological body close to the wound and the nerve network 20 can be less than the length of the wound. The less the distance between the neurons of the biological body close to the wound and the nerve network 20, the shorter the time of connecting the nerve network 20 and the neurons of the biological body, and the shorter the wound recovery time. In one embodiment, the area of the nerve graft is substantially parallel to a surface of the carbon nanotube structure 12, and the area of the nerve graft is greater than 15×15 square millimeters.

Referring to FIG. 9, a method for forming a nerve graft 100 includes the steps of:

S10, providing a carbon nanotube structure;

S20, forming a hydrophilic layer on a surface of the carbon nanotube structure;

S30, polarizing the hydrophilic layer to form a polar surface on the hydrophilic layer;

S40, seeding a number of neurons on the polar surface of the hydrophilic layer; and

S50, culturing the neurons until adjacent neurons connect with each other to form a nerve network.

In the step S10, the carbon nanotube structure has a number of carbon nanotubes capable of forming a free-standing structure. The free-standing structure of the carbon nanotube structure is realized by the carbon nanotubes joined by van der Waals attractive force. The carbon nanotube structure can include at least one carbon nanotube film. The carbon nanotube film can be a flocculated carbon nanotube film as shown in FIG. 2, a pressed carbon nanotube film as shown in FIG. 2, or a drawn carbon nanotube film as shown in FIG. 4. In addition, the carbon nanotube structure can include at least one carbon nanotube wire. The carbon nanotube wire can be an untwisted carbon nanotube wire as shown in FIG. 6 or a twisted carbon nanotube wire as shown in FIG. 7. In one embodiment, the carbon nanotube structure is formed by 30 layers of drawn carbon nanotube films. The angle between the aligned directions of the adjacent drawn carbon nanotube films is substantially 90 degrees.

In the step S20, the hydrophilic layer is formed on the surface of the carbon nanotube structure by evaporation or sputtering. The material of the hydrophilic layer is hydrophilic. For example, the hydrophilic layer is made from inorganic materials, such as silicon dioxide, titanium dioxide, iron oxide, or any combination thereof. In one embodiment, the carbon nanotube structure formed by 30 layers of drawn carbon nanotube films is fixed at a frame, and then a silicon dioxide layer is formed on the surface of the carbon nanotube structure by electron beam evaporation.

In the step 30, the polar surface can be formed on the hydrophilic layer by the steps of:

(a1), providing a supporter;

(b1), placing the carbon nanotube structure having the hydrophilic layer on a surface of the supporter; and

(c1), forming the polar surface on the hydrophilic layer by soaking the carbon nanotube structure having the hydrophilic layer located on the supporter.

In step (b1), the carbon nanotube structure can be located on part of the surface of the supporter. To decrease a specific surface area of the carbon nanotube structure and increase an adhesive attraction force between the carbon nanotube structure and the supporter, the step (b1) can further include the following steps: (b11), soaking the carbon nanotube structure located on the surface of the supporter with an organic solvent; and (b12), evaporating the organic solvent from the carbon nanotube structure. In one embodiment, the supporter is a plastic petri dish or an observation dish.

In step (c1), the carbon nanotube structure can be soaked with a polyamino acid solution or a polyetherimide solution to form the polar surface on the hydrophilic layer. For example, the polyamino acid solution or the polyetherimide solution can be sprayed on the surface of the hydrophilic layer. In one embodiment, to soak the hydrophilic layer with the polyamino acid solution or the polyetherimide solution, the step (c1) includes the following steps: (c11), dripping the polyamino acid solution or the polyetherimide solution on the surface of the hydrophilic layer; and (c12), purging the polyamino acid solution or the polyetherimide solution from the hydrophilic layer with deionized water after the polar surface is formed on the hydrophilic layer. In one embodiment, the polyamino acid solution is dripped on the carbon nanotube structure for about 10 hours. A concentration of the polyamino acid solution can be about 20 milligrams per milliliter.

In the step S30, the carbon nanotube structure can be further sterilized. Sterilizing the carbon nanotube structure kills all of the bacteria distributed in the carbon nanotube structure. The carbon nanotube structure can be sterilized by means of an ultraviolet sterilization technology or a high temperature sterilization technology.

In the step S40, the neurons can be from a mammal, such as a human, a mouse, or a cow. In one embodiment, the neurons are hippocampal neurons from a mouse. The neurons can be seeded on the hydrophilic layer by spraying a neuron solution to the polar surface of the hydrophilic layer, or by dipping the culture substrate into the neuron solution. In one embodiment, when the polar surface of the hydrophilic layer is covered by the neuron solution, the neurons in the neuron solution can be deposited or seeded on the polar surface of the hydrophilic layer.

In step S50, the conditions for culturing the neurons to branch and connect to other neurons are not limited. The neurons can be cultured under room temperature and standard atmospheric pressure. The neurons can also be cultured under a condition similar to a condition in a mammal. Referring to FIG. 10, when one of the neurons seeded on the polar surface of the hydrophilic layer are cultured in a typical clean room under room temperature and standard atmospheric conditions for about 15 days, a number of neurites branch from one of the neurons.

Referring to FIG. 11, if there are a number of neurons located one the polar surface of hydrophilic layer, the neurites from one neuron would preferentially extend to adjacent neurons. Thus, growth directions of the neurites can be guided by positions of the neurons. The neurons can also be connected by the neurtites to form the nerve network. Cell growth factors can be provided by the polar surface of the hydrophilic layer for culturing the neurons.

Referring to FIG. 12, one embodiment of a nerve graft 200 includes a culture substrate 30 and a nerve network 20 positioned on a surface of the culture substrate 30. The nerve network 20 includes a number of neurons 21. The adjacent neurons 21 connect with each other. The surface of the culture substrate 30 has a polarity attracted to a polarity of a surface of the nerve network 20 such that the nerve network 20 can closely adhere to the surface of the culture substrate 30.

The culture substrate 30 includes a carbon nanotube structure 12, a hydrophilic layer 14, and a polar layer 36. The hydrophilic layer 14 is located on a surface of the carbon nanotube structure 12. The polar layer 36 is located on a surface of the hydrophilic layer 14 away from the carbon nanotube structure 12. The nerve network 20 is positioned on a surface of the polar layer 36 away from the hydrophilic layer 14. The hydrophilic layer 14 is located between the carbon nanotube structure 12 and the polar layer 36.

The hydrophilic layer 14 makes the carbon nanotube structure 12 a hydrophilic environment for growing the neurons 21 of the nerve network 20. In addition, the hydrophilic layer 14 makes the polar layer 36 adhere easily to the carbon nanotube structure 12. The thickness of the hydrophilic layer 14 is in a range from about 1 nanometer (nm) to about 100 nm. Preferably, the hydrophilic layer 14 has a thickness in a range from about 1 nm to about 50 nm. The hydrophilic layer 14 is made from inorganic materials, such as silicon dioxide, titanium dioxide, iron oxide, or any combination thereof. In one embodiment, the hydrophilic layer 14 is a silicon dioxide layer with a thickness of about 10 nm.

The polar layer 36 is capable of providing a polarity such that the polarity of the culture substrate 30 can attract the surface of the nerve network 20. Thus, the culture substrate 30 is capable of providing a bio-compatible environment for seeding the neurons 21 and forming the nerve network 20. A material of the polar layer 36 can be polyamino acid, polyetherimide, or any combination thereof. The polyamino acid can be poly-D-lysine (PDL). In one embodiment, the polar layer 36 is a PDL layer.

Referring to FIG. 13, a method for forming a nerve graft 200 includes the steps of:

S11, providing a carbon nanotube structure;

S21, forming a hydrophilic layer on a surface of the carbon nanotube structure;

S31, forming a polar layer on a surface of the hydrophilic layer to form the culture substrate;

S41, seeding a number of neurons on a surface of the polar layer; and

S51, culturing the neurons until adjacent neurons connect with each other to form a nerve network.

In the step S11, the carbon nanotube structure has a number of carbon nanotubes capable of forming a free-standing structure. The free-standing structure of the carbon nanotube structure is realized by the carbon nanotubes joined by van der Waals attractive force. The carbon nanotube structure can include at least one carbon nanotube film. The carbon nanotube film can be a flocculated carbon nanotube film as shown in FIG. 2, a pressed carbon nanotube film as shown in FIG. 2, or a drawn carbon nanotube film as shown in FIG. 4. In addition, the carbon nanotube structure can include at least one carbon nanotube wire. The carbon nanotube wire can be an untwisted carbon nanotube wire as shown in FIG. 6 or a twisted carbon nanotube wire as shown in FIG. 7. In one embodiment, the carbon nanotube structure is formed by 30 layers of drawn carbon nanotube films. The angle between the aligned directions of the adjacent drawn carbon nanotube films is substantially 90 degrees.

In the step S21, the hydrophilic layer is formed on the surface of the carbon nanotube structure by evaporation or sputtering. The material of the hydrophilic layer is hydrophilic. For example, the hydrophilic layer is made from inorganic materials, such as silicon dioxide, titanium dioxide, iron oxide, or any combination thereof. In one embodiment, the carbon nanotube structure formed by 30 layers of drawn carbon nanotube films is fixed at a frame, and then a silicon dioxide layer is formed on the surface of the carbon nanotube structure by electron beam evaporation.

In the step S31, the polar layer can be formed on the surface of the hydrophilic layer by the steps of:

(a2), providing a supporter;

(b2), placing the carbon nanotube structure having the hydrophilic layer on a surface of the supporter; and

(c2), forming the polar layer on the carbon nanotube structure by soaking the carbon nanotube structure having the hydrophilic layer located on the supporter.

In step (b2), the carbon nanotube structure can be located on part of the surface of the supporter. To decrease a specific surface area of the carbon nanotube structure and increase an adhesive attraction force between the carbon nanotube structure and the supporter, the step (b2) can further include the following steps: (b21), soaking the carbon nanotube structure located on the surface of the supporter with an organic solvent; and (b22), evaporating the organic solvent out of the carbon nanotube structure. In one embodiment, the supporter is a plastic petri dish or an observation dish.

In step (c2), the carbon nanotube structure can be soaked with a polyamino acid solution or a polyetherimide solution to form the polar layer on the hydrophilic layer. For example, the polyamino acid solution or the polyetherimide solution can be sprayed on the surface of the hydrophilic layer. In one embodiment, to soak the hydrophilic layer with the polyamino acid solution or the polyetherimide solution, the step (c2) includes a step of dripping the polyamino acid solution or the polyetherimide solution on the hydrophilic layer located on the carbon nanotube structure. In one embodiment, the polyamino acid solution is dripped on the carbon nanotube structure for about 10 hours. A concentration of the polyamino acid solution can be about 20 milligrams per milliliter.

In the step S31, the carbon nanotube structure having the polar layer can be further sterilized. Sterilizing the carbon nanotube structure having the polar surface kills all of the bacteria distributed in the carbon nanotube structure. The carbon nanotube structure can be sterilized by means of an ultraviolet sterilization technology or a high temperature sterilization technology.

In the step S41, the neurons can be from a mammal, such as human, a mouse, or a cow. In one embodiment, the neurons are hippocampal neurons from a mouse. The neurons can be seeded on the polar layer by spraying a neuron solution to the surface of the polar layer, or by dipping the culture substrate into the neuron solution. In one embodiment, when the polar layer is covered by the neuron solution, the neurons in the neuron solution can be deposited or seeded on the surface of the polar layer.

In step S51, the conditions for culturing the neurons to branch and connect to other neurons are not limited. The neurons can be cultured under room temperature and standard atmospheric pressure. The neurons can also be cultured under a condition similar to a condition in a mammal.

Accordingly, the present disclosure is capable of providing a method for forming a nerve graft. The nerve graft including a carbon nanotube structure has the following benefits. First, the hydrophilic layer covers the surface of the carbon nanotube structure such that the nerve graft has good hydrophilic property. Second, the carbon nanotube structure is capable of accommodating many different shapes. Third, both of the hydrophilic layer and the carbon nanotube structure can have good tactility, and nonmetal and bio-compatible properties. Thus, the nerve graft including the hydrophilic layer and the carbon nanotube structure can be transplanted into a biological body and form a shape as desired.

It is to be understood that the above-described embodiments are intended to illustrate rather than limit the disclosure. Any elements described in accordance with any embodiments is understood that they can be used in addition or substituted in other embodiments. Embodiments can also be used together. Variations may be made to the embodiments without departing from the spirit of the disclosure. The above-described embodiments illustrate the scope of the disclosure but do not restrict the scope of the disclosure.

Depending on the embodiment, certain of the steps of methods described may be removed, others may be added, and the sequence of steps may be altered. It is also to be understood that the description and the claims drawn to a method may include some indication in reference to certain steps. However, the indication used is only to be viewed for identification purposes and not as a suggestion as to an order for the steps. 

What is claimed is:
 1. A method for forming a nerve graft, comprising: providing a carbon nanotube structure; evaporating or sputtering an inorganic material on a surface of the carbon nanotube structure to form a hydrophilic layer; soaking the hydrophilic layer with a polyamino acid solution or a polyetherimide solution to polarize a surface of the hydrophilic layer to a polar surface, wherein the soaking the hydrophilic layer with the polyamino acid solution or the polyetherimide solution comprises sub-steps of: providing a supporter; placing the carbon nanotube structure having the hydrophilic layer on a surface of the supporter; and soaking the hydrophilic layer on the carbon nanotube structure on the surface of the supporter with the polyamino acid solution or the polyetherimide solution to polarize a surface of the hydrophilic layer to a polar surface; and completely removing the polyamino acid solution or the polyetherimide solution from the polar surface; and seeding and culturing a plurality of neurons on the polar surface until adjacent neurons of the plurality of neurons are connected with each other, and the plurality of neurons form a nerve network.
 2. A method for forming a nerve graft, comprising: providing a culture substrate comprising a carbon nanotube film and a hydrophilic layer comprising an inorganic material located on the carbon nanotube film; soaking a surface of the hydrophilic layer of the culture substrate with a polyamino acid solution or a polyetherimide solution to polarize the surface of the hydrophilic layer to a polar surface, and then completely removing the polyamino acid solution or the polyetherimide solution with deionized water from the polar surface; and seeding and culturing a plurality of neurons on the polar surface until adjacent neurons of the plurality of the neurons are connected with each other, and the plurality of neurons form a nerve network.
 3. The method of claim 2, wherein the carbon nanotube film comprises a plurality of carbon nanotubes substantially parallel to a surface of the carbon nanotube film.
 4. The method of claim 3, wherein the inorganic material is selected from the group consisting of silicon dioxide, titanium dioxide, iron oxide, and any combination thereof.
 5. The method of claim 1, wherein the step of placing the carbon nanotube structure having the hydrophilic layer on the surface of the supporter further comprises: soaking the carbon nanotube structure on the supporter with an organic solvent; and evaporating the organic solvent from the carbon nanotube structure.
 6. The method of claim 1, wherein the step of soaking the hydrophilic layer with the polyamino acid solution or the polyetherimide solution comprises: dripping the polyamino acid solution or the polyetherimide solution on the surface of the hydrophilic layer on the supporter for about 10 hours to polarize the surface of the hydrophilic layer to the polar surface; and purging the polyamino acid solution or the polyetherimide solution from the hydrophilic layer with deionized water after the polar surface is formed on the hydrophilic layer.
 7. The method of claim 1, wherein a concentration of the polyamino acid solution is about 20 milligrams per milliliter.
 8. The method of claim 1, wherein the polyamino acid solution is a poly-D-lysine (PDL) solution.
 9. The method of claim 1, wherein the carbon nanotube structure comprises at least one carbon nanotube film comprising a plurality of carbon nanotubes substantially parallel to a surface of the at least one carbon nanotube film.
 10. The method of claim 9, wherein each of the at least one carbon nanotube film is a flocculated carbon nanotube film, a pressed carbon nanotube film, or a drawn carbon nanotube film.
 11. The method of claim 9, wherein the carbon nanotube structure comprises a plurality of carbon nanotube films stacked together, and adjacent carbon nanotube films of the plurality of carbon nanotube films are combined and attracted to each other only by van der Waals attractive force therebetween.
 12. The method of claim 1, wherein the carbon nanotube structure comprises at least one untwisted carbon nanotube wire.
 13. The method of claim 1, wherein a thickness of the hydrophilic layer is in a range from about 1 nanometer to about 100 nanometers. 